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Researchers at The University of Texas MD Anderson Cancer Center have determined why many cancer vaccines fail to elicit a
robust T cell response-the cells hover near the vaccination site instead of
migrating to the tumor.1 The surprisingly simple solution is
switching the vaccine formulation to a different carrier.

Vaccination with peptide antigens typically elicits both
antibody- and cell-based immune responses that are mediated by B cells and T
cells, respectively. A robust response by CD8+ cytotoxic T cells is especially
important in fighting tumors.

Ideally, a tumor-derived antigen vaccine would cause
antigen-presenting cells (APCs) such as dendritic cells (DCs) to pick up the
antigen and carry it to lymph nodes, where they would then activate
B cells that make tumor-targeting antibodies and CD8+ cytotoxic T cells that hunt tumor
cells.

In practice, raising antibodies against tumor antigens is
easy, but getting a robust T cell response has proven challenging.

The only marketed cancer vaccine in the U.S.-Dendreon Corp.'s Provenge sipuleucel-T for
prostate cancer-takes a shortcut to eliciting a strong T cell response.
Provenge therapy involves extracting and activating patient-derived DCs ex
vivo, then reintroducing the DCs into patients in large enough numbers to
promote tumor-targeting T cell activity.

Direct in vivo vaccination could help avoid the cost
and complexity of autologous cell therapies like Provenge but would only be
effective if a stronger
T cell response could somehow be elicited.

He said prior preclinical and clinical studies have shown
that cancer vaccines typically cause an initial burst of T cell activity, but
the response quickly peters out.

Failure to launch

Overwijk's team vaccinated a mouse model
for melanoma with a short fragment of the melanoma antigen silver homolog (SILV;
PMEL17;
GP100)
in a solution of incomplete Freund's
adjuvant (IFA),
a commonly used mineral oil-based carrier.

As expected, the formulation elicited a strong B cell
response but only a weak T cell response.

To understand why the vaccine-induced T cells were not very
effective, Overwijk's team used a luciferase-based
imaging system to look at the localization of T cells in tumor-bearing mice.
Instead of migrating to the tumors, the antigen-specific T cells lingered
around the vaccine injection site for weeks.

"We saw that T cells were getting hung up at the
injection site," said Overwijk. "They expanded in number and hung out
in the blood near the injection site. Very few of them went to the tumor sites."

Overwijk found that IFA did not readily disperse in the
bloodstream and instead formed a long-lived depot of antigen that trapped T
cells. Instead of homing to tumor cells, the T cells were sequestered near the
vaccination site, formed a dense granuloma-like mass, became inactive and
eventually underwent apoptosis.

"We think it's the long persistence of the vaccine
that leads to this effect. The mineral oil is not biodegradable," he
noted.

To overcome this problem, Overwijk's team experimented with
various vaccine formulations that are shorter lived than IFA. The team settled
on a formulation of GP100 and a cocktail of adjuvants suspended in a saline
buffer.

With the new saline-based formulation, "the vaccine
site becomes cleared of antigen in 4-5 days rather than 90 days with IFA,"
said Overwijk.

Mice vaccinated with the saline-based formulation showed T
cell migration away from the vaccination site and toward the tumor (see"Improving tumor vaccine
formulation"). The result was greater tumor-specific T
cell responses at the tumor site than those seen when using IFA-based controls.

Benjamin Chen, executive chairman of Immune Targeting Systems Ltd., said Overwijk's findings are potentially relevant to a
variety of
T cell-targeted vaccines for cancer and possibly other indications.

"It's been known that the IFA-like adjuvant used in
humans can cause granulomas and necrosis at injection sites," but the
significance of these structures was unclear, said Chen. "The phenomenon
of getting T cell infiltration and sequestration at the vaccination site and
absence of T cells at the tumors is a very important finding. We now know the
problem, and there's now a hypothesis to address this."

Immune Targeting is developing self-adjuvanting, T cell-targeted
peptide vaccines. A Phase IIa trial of the company's most advanced product-Flunisyn influenza A vaccine
(FP-01.1)-met
its primary endpoint of T cell induction. The company also has preclinical
vaccine candidates for HBV and an undisclosed cancer indication.

Cornelis Melief, CSO of vaccine maker ISA Pharmaceuticals B.V.,
suspects that the carrier may not be the only culprit that hinders T cell
migration. He noted that the poor efficacy of the mineral oil-based vaccine in
the paper could result from the excessively short peptides used in the study.

ISA's vaccines use long peptides in an IFA-like carrier.
The company's ISA-HPV-01, a therapeutic
vaccine for HPV, is expected to start
Phase II testing this year for advanced cervical cancer and is in Phase I/II
trials for anal intraepithelial neoplasia.

Melief said the short GP100 antigen peptide used by
Overwijk's team binds to MHC I receptors found
on a multitude of cells, including the T cells themselves. As a result,
vaccination with this short peptide can overstimulate the T cells and
discourage proper presentation of antigens by DCs and other APCs.

Many researchers prefer using short-peptide antigens
because they are easier to work with than longer peptides and do not require
additional processing by APCs.

Melief believes a better approach is to use long-peptide
antigens. Long peptides do not readily bind to MHC I and can only be taken up
by DCs, which are less numerous than other cell types, including T cells.
Instead of the excessive local response caused by short peptides, long peptides
are carried from the injection site by DCs to lymph nodes, leading to a more
measured and productive immune response.

"MHC I-binding peptides are lousy vaccines because
they exogenously load onto all cells with class I MHC," said Melief. "This
leads to a tolerizing mode of vaccination. If you are going to deliver short
peptides, Overwijk clearly shows that the only way is to provide them along
with strong adjuvants and saline."

Last year, Melief and collaborators at the Leiden University Medical Center
reported that long-peptide vaccines induce an effective antitumor T cell
response and tumor regression in a mouse model for HPV even when formulated
with an IFA-like carrier.2

Likewise, Melief noted that the supplementary data in
Overwijk's paper suggest that when the team used longer peptides, the melanoma
vaccine appeared to work well even in IFA.

Overwijk said Melief is correct that "when given in
IFA, long peptides are superior to short peptides, possibly because the
intensity of antigen presentation of long peptides is lower."

However, he said, "we still see eventual T cell
dysfunction after vaccination with long peptides in IFA, probably since the
persistence of antigen is still too long."

Chen said the challenge now for vaccine makers is to
maximize the exposure of antigens to APCs without causing long-term T cell
sequestration. He said optimal local persistence of the antigen may vary from
one vaccine to another.

"You want to attract antigen-presenting cells to come
in and pick up the antigen and prime T cells, but you don't want the depot to
be around for too long," said Chen. "You need to make sure the T
cells get primed and home to where they are needed as quickly as possible."

Overwijk said a saline-based carrier is probably too
short-lived to produce optimal DC responses. His team is now testing a variety
of water-soluble carriers and materials including gels, microsomes and
nanoparticles. The goal is to deliver the antigen in a way that mimics acute
viral infection, which is familiar territory for the immune system.

"We're currently working with industry and academia
to try a variety of particle-based, intermediate-release formulations,"
said Overwijk. "We haven't found the ideal formulation yet, but we think
we want it to be around for around five days, which is about as long as a virus
hangs around."

Overwijk did not disclose the companies with whom he is
working. He did not patent his findings.

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